title: Generation and evaluation of a human corneal model cell system for ophthalmologic issues using the HPV16 E6/E7 oncogenes as uniform immortalization platform.
abstract: The present study aimed at employing the human papillomavirus type 16 (HPV16) E6/E7 gene platform, to create a uniform authentic in vitro model cell system of the human cornea for ophthalmologic issues and here especially for prospective biomaterial evaluations for therapeutic regenerative approaches. Therefore, HPV16 E6/E7 genes were employed as uniform platform to immortalize primary human corneal keratinocytes (IHCK), fibroblasts (IHCF), and endothelial (IHCE) cells. qPCR revealed that E6/E7 mRNA transcription persisted at rising passages and FISH detection of the chromosome portfolio 1, 8, 10 and 18 showed fairly the disomic cytogenetic status. Hot spot passages proved oscillation of aneuploidies in the entire passage spectrum under study, while hot spot aneuploidies annotated prevalence for distinct chromosomes. Though IIF revealed general endurance, tissue-innate corneal biomarkers were modulated, i.e. expressed in a temporal-confluence, temporal-spatial or passage-dependent manner. In summary, by the fairly normal chromosomal status, and expression of tissue-innate biomarkers, we created for the first time a uniform authentic in vitro model cell system of the human cornea, by application of the HPV16 E6/E7 immortalization platform only. This system renders a precious tool for prospective iterative in vitro studies on issues such as corneal tissue homeostasis, pharmaceutical generics, and/or evaluation of new biomaterials for clinical corneal applications.
authors.affiliation: Department of Oral Biotechnology, Dental School, University Medical Center Freiburg, Hugstetterstrasse 55, 79106 Freiburg, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Schulz
authors.forename: Thorsten
authors.initials: T
authors.lastname: Steinberg
authors.forename: David
authors.initials: D
authors.lastname: Beck
authors.forename: Pascal
authors.initials: P
authors.lastname: Tomakidi
authors.forename: Rosita
authors.initials: R
authors.lastname: Accardi
authors.forename: Massimo
authors.initials: M
authors.lastname: Tommasino
authors.forename: Thomas
authors.initials: T
authors.lastname: Reinhard
authors.forename: Philipp
authors.initials: P
authors.lastname: Eberwein
mesh_terms: D002455:Cell Division / Q000502:physiology; D002472:Cell Transformation, Viral / Q000235:genetics; D002478:Cells, Cultured; D003315:Cornea / Q000166:cytology* / Q000378:metabolism / Q000821:virology; D052162:Human papillomavirus 16 / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism; D008297:Male; D050725:Papillomavirus E7 Proteins / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D050725:Papillomavirus E7 Proteins
keywords: Corneal model cell system; Homeostasis; Human cornea; Human papillomavirus type 16; Immortalization